Glycoside Hydrolase Family 62

This page has been approved by the Responsible Curator as essentially complete. CAZypedia is a living document, so further improvement of this page is still possible. If you would like to suggest an addition or correction, please contact the page's Responsible Curator directly by e-mail.

Glycoside Hydrolase Family GH62
Clan	GH-F
Mechanism	inverting
Active site residues	Known
CAZy DB link
https://www.cazy.org/GH62.html

Substrate specificities

This small family of glycoside hydrolases comprises both eukaryotic and prokaryotic enzymes. All the characterized enzymes in this family are arabinofuranosidases and the majority act on xylose moieties in xylan and arabinose moieties in arabinan that are single substituted with α-1,2 and α-1,3-L-arabinofuranose side chains [1] with K_cat ranging from 0.3 to 180 s^-1 on wheat arabinoxylan [2, 3, 4]. Interestlingly, the preference for α-1,2 and α-1,3-L-arabinofuranose side chains varies for GH62s, hence the catalytic rate for the two side chains vary[5, 6]. However, a single GH62 enzyme from Pencillium oxalicum exclusively act on the α-1,3-L-arabinofuranose side chains [7]. The GH62 enzymes also display limited non-specific arabinofuranosidase activity; for example the arabinofuranosidases exhibit no [8] or very little [2, 3] activity against 4-nitrophenyl α-L-arabinofuranoside. Several of these enzymes contain carbohydrate binding modules that target cellulose[8] or xylan[9].

Kinetics and Mechanism

The stereochemical course of arabinose was followed by ¹H NMR during hydrolysis of a 50:50 mixture of XA²XX:XA³XX by Aspergillus nidulans α-L-arabinofuranosidase A, resulting in the release of β-furanose demonstrating that GH62 enzymes in fact are inverting enzymes [4], which is in accordance with the known inverting mechanism for GH43 [10] constituting clan F with GH62 [11]. Due to arabinose's fast mutarotation, however, the anomeric signal decreased considerably already after 1 min, which was overcome by recording the first spectrum 23 s after enzyme addition [4].

Catalytic Residues

Asp (general base) and Glu (general acid), as suggested by tertiary structures [2, 3, 12] and supported by site-directed mutagenesis and kinetic data [2, 3].

Three-dimensional structures

Based on its location in clan F together with GH43, enzymes from family GH62s were predicted to display a 5-fold β-propeller fold. This hypothesis was confirmed by three papers published in 2014 [2, 3, 12]. The predicted catalytic general acid, catalytic general base and pKa modulator [13] were also confirmed by mutagenesis data [2, 3]. The active site arabinose-containing pocket opens up into a cleft or channel that binds the xylooligosaccharides and thus the xylan chain. The residues that interact with the substrate backbone were identified for Streptomyces coelicolor α-L-arabinofuranosidase A (ScAbf62A) in a crystal structure in complex with xylopentaose, which spanned subsite +2R to +4NR [2]. In this respect a conserved tyrosine, present on a mobile loop, was shown to make an important contribution to substrate binding through hydrophobic interactions with the arabinose located in the active site [14]. Remarkably, the xylan main chain bound in two orientations in the crystal structures of ScAbf62A and Streptomyces thermoviolaceus α-L-arabinofuranosidase A, as may be required to position both single α-1,2 and α-1,3-L-arabinofuranose side chains in subsite -1 for productive binding in the active site pocket [2, 3]. The preference for either α-1,2 or α-1,3-L-arabinofuranose side chains seems to correlate with the presence of an arginine residue interacting with the xylan main chain at the +2R subsite [6].

Family Firsts

First sterochemistry determination: Determined for Aspergillus nidulans α-L-arabinofuranosidase A by ¹H NMR [4].
First general acid residue identification: 3D structural data [2, 3, 12] in concert with supporting mutagenesis data [2, 3].
First general base residue identification: 3D structural data [2, 3, 12] in concert with supporting mutagenesis data [2, 3].
First 3-D structure: Several papers in 2014 reveal the 5-fold β-propeller fold [2, 3, 12].

References

All Medline abstracts: PubMed